Role of temperature in regulation of ovarian cycle in bull frog Rana tigerina

Effect of temperature on the ovarian cycle was studied in R. tigerina by exposing them to (1) constant low (22 degrees C) temperature during preparatory (active vitellogenic growth) phase (March-May) when the mean ambient temperature ranged from 26 degrees-28 degrees C and (2) to constant high (30 degrees +/- 1 degrees C) temperature during postbreeding regression phase (August-November) when the mean ambient temperature ranged from 22 degrees-26 degrees C. The ovaries of initial controls (biopsy samples taken prior to the commencement of the experiment) in March contained only first growth phase (FGP) oocytes with a maximum size range of 361-480 microns in diameter. In the frogs exposed to constant low temperature for 2 months, only 7% of FGP oocytes were recruited to second growth phase (SGP) with a mean largest diameter of 631 microns compared to 31% large SGP oocytes with a mean diameter of 1114 microns in the frogs collected from natural fields. The number of atretic follicles (AF) was lower and fat body weights were significantly higher in low temperature exposed frogs. The exposure of the frogs to constant high temperature during postbreeding months caused an increase in the mean diameter and number of large FGP oocytes, numerical increase in AF and decrease in fat body weights over corresponding controls maintained at room temperature. The pituitary gonadotrophs of these frogs showed stimulatory changes such as increase in cell size and appearance of secretory granules in the cytoplasm. The results suggest that in R. tigerina high temperature stimulates oocyte growth while low temperature retards it.(ABSTRACT TRUNCATED AT 250 WORDS)     

An ultrastructural study of the spleen of the ranid frog Rana perezi

The spleen of Rana perezi is encapsulated by connective tissue and shows by light microscopy two areas with no obvious border: the white pulp and the red pulp. The white pulp-lymphoid clusters are scattered throughout the organ and contain lymphocytes, reticular cells, and some plasma cells. The red pulp displays two different portions. The predominant region consists of reticular cells, lymphocytes, a variety of other leucocytes, and cells undergoing division. This area possibly performs a haemopoietic function. The smaller portion of the red pulp is characterized by reticular-phagocytic cells and may be haemocaretic in its function. Macrophages and pigmented cells occur in both white and red pulp. The organization of the spleen of R. perezi can be considered as a transitional or intermediate state between the primitive condition seen in certain fishes and amphibians and the more complex organ of ammiotes.     

Type-1 cannabinoid receptor expression in the frog, Rana esculenta, tissues: a possible involvement in the regulation of testicular activity

Endogenous cannabinoids and type-1 cannabinoid receptor (CB1) are widely produced and distributed in the central nervous system (CNS) and peripheral nerves in mammals. In addition, the detection of endocannabinoids and corresponding receptors in non nervous peripheral tissues indicates an involvement of the system in the control of a wide range of physiological activities, including reproduction. Recently, the existence of CB1 was also observed in lower vertebrates and in urochordate suggesting that the endocannabinoid system is phylogenetically conserved. Using RT-PCR, CB1 mRNA expression profiles were characterized in a wide range of tissues of the anuran amphibian, the frog, Rana esculenta. Besides a strong expression in the CNS, CB1 was also present in testis, kidney, liver, ovary, muscle, heart, spleen, and pituitary. The CB1 expression pattern has been characterized in both testis and CNS during the annual sexual cycle. In testis, CB1 is poorly expressed during the winter stasis of the spermatogenesis rising during the breeding season and resumption period. An expression profile mismatching to that observed in testis was detected in whole-brain preparations during the sexual cycle; in particular in the diencephalon, the encephalic area mainly involved in the control of reproductive functions. Furthermore, fluctuations inside isolated encephalic areas and spinal cord were observed all over the reproductive cycle. In conclusion, CB1 receptor is expressed in R. esculenta CNS and testis. As far as the gonad it concerns, our results suggest the involvement of the endocannabinoids in the control of reproductive function.     

Role of photoperiod and temperature in seasonal morph determination of the butterfly Eurema hecabe

ABSTRACT. Environmental cues affecting seasonal morph determination of the pierid butterfly Eurema hecabe L. were examined in laboratory experiments and in the field. A sexual difference exists in the photoperiodic response for seasonal morph determination: in short days at 25C the proportion of the autumn morph is higher in females than in males, and as the rearing temperature is decreased the proportion of autumn morphs increases in both sexes. A 5C drop in temperature during the final larval stadium induces 100% autumn morph production in short-day animals. This increase in autumn morph production induced by temperature decrease explains the seasonal morph change of this butterfly occurring in early autumn in the field. The role of this sex-linked photoperiodic response is discussed as a seasonal strategy of reproduction.     

Activation of AMP-dependent protein kinase by hypoxia and hypothermia in the liver of frog Rana perezi

We have investigated different signaling molecules that could be activated by temperature acclimation and hypoxia, using an experimental approach consisting in submerging frogs in a water-filled box maintained at 2-4 degrees C at ambient oxygen levels or supplied with 98% N2:2% CO2 for normoxia or hypoxia conditions, respectively. The results obtained showed no significant changes in the expression of heat shock protein 70. The phosphorylation state of AMP-dependent activated protein kinase, the down-stream component of a protein kinase cascade that acts as an intracellular energy sensor, was significantly increased in both experimental conditions, showing higher values in the absence of oxygen. Similarly, the phosphorylation state of one of its known substrates, elongation factor 2, was also increased, consistent with the arrest of protein synthesis. These results point out an important role of this kinase, adjusting the rates of ATP-consuming and ATP-generating pathways, in the survival strategies to hypoxia and hypothermia.     

Histology of the esophagus of the adult frog Rana perezi (Anura: Ranidae).

Study of the esophageal microscopic morphology of adult Rana perezi by light and electron microscopy discloses some large folds throughout the esophagus that are in themselves ringed. Glandular ostia open in the furrows of the luminal surface. The esophageal wall is made up of a connective adventitia rich in melanocytes, a muscular tunica, a connective and glandular subepithelial layer, and a pseudostratified ciliated epithelium. This epithelium basically consists of ciliated, goblet, basal, microvillous-apex, and migratory cells. Two types of goblet cells are distinguished with regard to the granular ultrastructure. The microvillous-apex cell has not been found in other amphibians. It shows a very differentiated morphology with a high number of mitochondria. The basal cells give the epithelium a pseudostratified morphology, and they have a proliferative function. Glands are branched and drain through an excretory duct that has a monolayered mucosecreting epithelium. The glandular units are formed by two principal types of cells: mucosecretory and serous.     

Ovarian activity and reproduction in the frog, Rana esculenta

Rana esculenta specimens were collected, during the last 13 years, in well-defined areas around Naples. The annual ovarian cycle shows distinct phases of recrudescence (starting September; vitellogenesis), breeding (late March-early July; egg deposition and active oogenesis) and quiescence (July-August; no follicular growth). Previtellogenic follicles are recruited for vitellogenesis in early September and in between two successive ovulatory waves. Breeding congregations are generally formed after a heavy rain fall and eggs are laid in standing waters, temporary or permanent. A maximum of three clutches of eggs is produced during the breeding season, at roughly monthly intervals. All mature females reproduce to some extent. Ovarian weight and clutch size are positively correlated to body weight. Depending upon the body size, the potential clutch size ranges from 1000 to 3500 eggs during the first wave of ovulation and it is notably smaller in the successive wave(s) of ovulation. Egg masses and tadpoles are left unprotected and mortality is high. The life cycle from the fertilized egg to completion of metamorphosis is 2 months and oogenesis in the ovary starts in the larva before the onset of metamorphic climax. Young females hatching from the first clutch of eggs may reach sexual maturity and breed in May the following year; those hatching from the last clutch require nearly 20 months to reach sexual maturity. The importance of some endocrine and exocrine factors for the regulation of ovarian activity and reproduction is discussed.     

The subcommissural organ of the frog Rana perezi is innervated by nerve fibres containing GABA

The innervation of the frog subcommissural organ was studied by light-microscopic and ultrastructural immunocytochemistry using antisera against serotonin, noradrenaline, dopamine, gamma-aminobutyric acid (GABA), glutamic acid decarboxylase, different GABA receptor subunits and bovine Reissner's fibre material (AFRU). In the proximity of the organ, serotonin- and noradrenaline-containing fibres were rare whereas dopamine-immunoreactive fibres were more numerous. Many GABA- and glutamic acid decarboxylase-containing nerve fibres were found at the basal portion of the ependymal cells of the subcommissural organ. Under the electron microscope, these GABA-immunolabelled nerve endings appeared to establish axoglandular synapses with secretory ependymal cells of the subcommissural organ. In addition, the secretory ependymal cells expressed high amounts of the beta2-subunit of the GABA(A) receptor. Since GABA-immunoreactive neurons were present in the frog pineal organ proper and apparently contributed axons to the pineal tract, we suggest that at least part of the GABAergic fibres innervating the frog subcommissural organ could originate from the pineal organ.     

The influence of environmental temperature and photoperiod on activity in the red squirrel,Tamiasciurus hudsonicus

The effects of environmental temperature and photoperiod on wheelrunning activity were studied in the diurnal red squirrel,Tamiasciurus hudsonicus. From 1 July to 22 November 1973 eight squirrels were maintained in separate cages in an outdoor shelter. Activity usually began before sunrise in the summer an early fall and after sunrise in the late fall. Wheel-running usually persisted throughout the day and ceased about the time of sunset in the summer and early fall. In late fall cessation frequently occurred after sunset. Neither changes in temperature nor photoperiod were found to separately account for the seasonal changes. Multiparameter analysis of variance was found to be a powerful statistical tool for analyzing the data. Six models for predicting the onset of activity were constructed and tested.This publication is based on a Master's thesis by B. R. Bahnak.     

Erythrocytic organic phosphates: diel and seasonal cycles in the frog, Rana berlandieri

Hemoglobin (Hb) concentration, hematocrit (Hct), total organic phosphates (Ptot), nucleoside triphosphates (NTP), 2,3-diphosphoglycerate (DPG), and "inositol polyphosphates" (IPP) were measured in the erythrocytes of adult frogs at different times of day in winter and summer after acclimatization to 15 degrees C and an LD 12:12 photoperiod. The same measurements were also made on animals acclimated to LD 16:8 in summer. All of the measured parameters varied significantly at different times of the day and between seasons in animals acclimated to an LD 12:12 photoperiod. Summer animals acclimated to LD 16:8 showed significant diel cycles only in Hct and Hb.     

Neural organization of the ventilatory activity in the frog,Rana catesbeiana. II

The paralyzed, decerebrate frog, Rana catesbeiana, displays “fictive” oropharyngeal and pulmonary ventilations. In order to evaluate the neuronal correlates of these two centrally programmed ventilatory bursting patterns, we have performed intra-and extracellular recordings of bulbar respiratory neurons in this fictively breathing preparation. A total of 123 respiratory neurons were recorded from the caudal medulla. Of 51 antidromically activated neurons, 20 were vagal motoneurons and 31 were hypoglossal motoneurons. Respiratory neurons that depolarized during the lung (L) or non-lung (N) ventilatory phases were classified as L or N neurons, respectively. Phase spanning neurons (S) were active during both L and N phases. Some neurons showed oscillations of membrane potential synchronous with oropharyngeal ventilation. Those active during the buccal elevation phase were exclusively L neurons whereas those having buccal depressor activity were exclusively N neurons. Synaptic drive potentials were observed in all neurons recorded intracellularly. In some neurons, hyperpolarization was caused by inhibitory postsynaptic potentials, as demonstrated by reversal of membrane potential trajectory after intracellular chloride iontophoresis. Some individual motoneurons and interneurons exhibited both pulmonary and buccal ventilatory activity, indicating that both pattern generators project to a common motor control system. 1994 John Wiley & Sons, Inc.     

Neural organization of the ventilatory activity in the frog,Rana catesbeiana. I

In order to elucidate the neural basis for lung ventilation in the frog, we have investigated the efferent neural activity to oropharyngeal muscles in the decerebrate, paralyzed, unanesthetized bullfrog, Rana catesbeiana. Efferent motor output was recorded from the mandibular branch of the trigeminal (Vmd), the laryngeal branch of the vagus (Xl), and the main and sternohyoid branches of the hypoglossal nerve (Hm and Hsh, respectively). Two types of rhythmic bursting outputs were observed: (1) a high-frequency, low-amplitude, reciprocal oscillation between Vmd, a buccal levator nerve, and Hsh, a buccal depressor nerve; and (2) a low-frequency, high amplitude, synchronous bursting of Vmd, Hm, Hsh, and Xl. The first type is inferred to represent fictive oropharyngeal ventilation. The second type of burst was divided into four intervals: (a)augmenting acitivity of Hsh; (b) activation of Xl with continued activation of Hsh; (c) activation of Vmd and Hm (a buccal levator nerve), continued activation of Xl, and termination of Hsh activity; and (d) waning activity in Vmd and Hm associated with a prominent second wave in Xl. This coordinated activity is inferred to represent fictive pulmonary ventilation because the neurograms in these four intervals correspond closely EMGs and neurograms recorded in the intact frog during the four phases of pulmonary ventilation, namely, buccal depression, pulmonary expiration, pulmonary inspiration, and glottal closure. Hypercapnia, vagotomy, and cutaneus pinching enhanced the high-amplitude, low-frequency rhythm, but not the low-amplitude, high-frequency oscillation. Lung inflation generally inhibited the former and not the latter, but in some cases lung inflation stimulated pulmonary ventilation. We conclude that oropharyngeal and pulmonary ventilation of the frog are produced by one or, possibly, two intrinsically active generators. 1994 John Wiley & Sons, Inc.     

Role of various regulation factors of interrenal gland activity in response to stress in the green frog Rana esculenta L]

The effects of an ether stress on the corticosterone levels were studied in variously experimented frogs. The removal of the distal lobe of the pituitary gland, with preservation of the intermediate lobe which contains also ACTH, suppresses, as total hypophysectomy, the increase of corticosterone level observed in normal frogs. The administration of dexamethasone reduces but not suppresses this increase. A second ether stress 20 minutes after the first is effective. The injection of frog plasma treated or not with an corticosteroid adsorbant do not alter the response to stress. These results suggest that modifications of corticosterone level, in some limits, do not disturb the effects of a stress.     

Glucose and caffeine regulation of liver glycogen phosphorylase activity in the freeze-tolerant wood frog Rana sylvatica

We have examined the effect of glucose and caffeine inhibition on the activity of liver glycogen phosphorylase a from the freeze-tolerant frog Rana sylvatica. Kinetic studies indicate that this enzyme exhibits similar sensitivity to glucose inhibition (glucose dissociation constant = 12.5 mM) as the mammalian enzyme. Little inhibition (less than 25%) was observed at normal glucose concentrations (1-5 mM), while significant inhibition (60-95%) occurred at glucose concentrations (50-500 mM) present in freezing-exposed animals. These results favour the hypothesis that in the normal state glucose regulates phosphorylase activity primarily through the promotion of dephosphorylation of phosphorylase a, whereas during freezing regulation is achieved through phosphorylase a inactivation. The caffeine dissociation constant (0.93 mM) and the degree of synergism between caffeine and glucose (interaction factor, alpha = 0.14) were also similar to that observed for the mammalian enzyme. Hence, if a caffeine-like ligand exists in vivo, it must be in low enough amounts during freezing to allow sufficient phosphorylase a activity for high glucose production.     

Effects of photoperiod and temperature on testicular function in amphibians

Most amphibians present an annual testicular cycle characterized by a quiescent period (late autumn-winter) and a spermatogenic period (spring and summer). At the end of the period of spermatogenesis undifferentiated interstitial cells transform into steroid-secreting Leydig cells which regress in spring at the beginning of the new spermatogenetic cycle. The testicular cycle is controlled by the pituitary gonadotropin levels which are high in autumn and winter, low in spring and increase temporarily in the middle of summer. Photoperiod and temperature seem to be the most important external factors involved in the regulation of this cycle in many amphibian species since the colder the geographic area, the longer the quiescent period and the shorter the spermatogenic period. This suggests the occurrence of a potentially continuous cycle in these species, in contrast with that which occurs in other species having an endogenous rhythm of testicular function which is much less sensitive to environmental factors. Although the specific response to temperature can vary widely between species, the most frequent observation in amphibians with a potentially continuous cycle is that exposure to mild temperatures (15-20 degrees C, according to the spring temperatures of the different geographic areas) stimulates spermatogenesis even during the period of testicular quiescence. If this mild temperature is combined with a long photoperiod, complete spermatogenesis is attained. Experiments performed during the period of germ-cell proliferation (development from spermatogonia to round spermatids) indicated that low temperatures (below 11 degrees C) as well as short photoperiods (less than 8 h of light) hinder germ-cell proliferation. Moderately high temperatures (about 30 degrees C) do not impair this proliferation. In the newt Triturus marmoratus, it has been shown that an excessively long photoperiod (over 16 h) has the same effect as a short photoperiod. In this species eyes are not required for the testicular photoperiodic response. Photoperiod appears to have no effect on spermiogenesis (differentiation of round spermatids into spermatozoa), because once round spermatids are formed, spermiogenesis will occur even in total darkness. Mild temperatures seem to be necessary for spermiogenesis as well as for androgen biosynthesis because neither process will take place at extreme temperatures. Results on the effect of photoperiod in steroidogenesis differ between species.